1. Field of the Invention
The availability of molecules which are able to specifically bind to a particular spatial and polar organization is the basis for a wide variety of techniques referred to as competitive protein binding assays. These techniques depend upon having a member of a specific binding pair conjugated with the label which is involved with the production of a detectible signal.
Depending upon the nature of the label, various methods have evolved for distinguishing between an analyte which is bound to the corresponding member of the specific binding pair and analyte which is unbound. With the various techniques, either the receptor or the ligand is labeled. Particularly, where the receptor is labeled, the receptor is normally one component of a complex mixture of analogous composition and molecular weight. For example, antibodies which are isolated from serum will be present with globulins and other antibodies which are either non-specific or specific for a wide variety of ligands other than the ligand of interest. When labeling the receptor composition, both the receptor of interest as well as the contaminating globulins will be labeled. The label involved with extraneous receptors or other materials will act as a background in the assay, interfering with the sensitivity of the assay.
While affinity chromatography may be employed to enhance the purity of the receptor of interest, this technique has many deficiencies. One deficiency is that the most strongly binding antibodies tend to be retained by the affinity chromatography column. Secondly, there are normally substantial losses of the antibodies of interest and substantial reduction in the binding constant of the recovered antibody. It is therefore desirable to find alternative methods to provide labeled reagents having reduced amounts of label bound to extraneous materials.
2. Brief Description of the Prior Art
U.S. Pat. No. 3,998,943 discloses a fluorescent immunoassay involving a ligand conjugated to a fluorescer, employing receptor to ligand and receptor to fluorescer, where the receptor to fluorescer is inhibited from binding to fluorescer when receptor to ligand is bound to ligand. U.S. Pat. No. 3,935,074 describes an immunoassay where a receptor for a detector label and a receptor for ligand are employed with the detector label labeled to ligand. Various labels are described. U.S. Pat. No. 3,996,345 describes an assay employing a chromogenic pair, where one of the chromogens fluoresces emitting light at a wavelength within the absorption band of the other chromogen. Copending patent application Ser. No. 815,636, filed July 14, 1977 now U.S. Pat. No. 4,160,145, discloses the use of a non-enzymatic catalyst as a label in competitive protein binding assays.
Co-pending application Ser. No. 893,910, filed Apr. 5, 1978, describes a chemiluminescent label in a competitive protein binding assay. Assays dependent upon the presence of enzyme labile bonds are described in Carrico, et al, Anal. Biochem. 72, 271-282 (1976); ibid 72, 283-292 (1972).